Cotranscriptional Folding of a 5′ Stem-loop in the Escherichia coli tbpA Riboswitch at Single-nucleotide Resolution

IF 4.7 2区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY
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引用次数: 0

Abstract

Transcription elongation is one of the most important processes in the cell. During RNA polymerase elongation, the folding of nascent transcripts plays crucial roles in the genetic decision. Bacterial riboswitches are prime examples of RNA regulators that control gene expression by altering their structure upon metabolite sensing. It was previously revealed that the thiamin pyrophosphate-sensing tbpA riboswitch in Escherichia coli cotranscriptionally adopts three main structures leading to metabolite sensing. Here, using single-molecule FRET, we characterize the transition in which the first nascent structure, a 5′ stem-loop, is unfolded during transcription elongation to form the ligand-binding competent structure. Our results suggest that the structural transition occurs in a relatively abrupt manner, i.e., within a 1–2 nucleotide window. Furthermore, a highly dynamic structural exchange is observed, indicating that riboswitch transcripts perform rapid sampling of nascent co-occurring structures. We also observe that the presence of the RNAP stabilizes the 5′ stem-loop along the elongation process, consistent with RNAP interacting with the 5′ stem-loop. Our study emphasizes the role of early folding stem-loop structures in the cotranscriptional formation of complex RNA molecules involved in genetic regulation.

Abstract Image

以单核苷酸分辨率观察大肠杆菌 tbpA 核糖开关中 5-́ 干环的同源折叠。
转录延伸是细胞中最重要的过程之一。在 RNA 聚合酶延伸过程中,新生转录本的折叠在基因决定中起着至关重要的作用。细菌核糖开关是 RNA 调节器的典型例子,它们通过改变代谢物感应时的结构来控制基因表达。之前的研究发现,大肠杆菌中的焦磷酸硫胺素感应 tbpA 核糖开关在同转录过程中会采用三种主要结构来实现代谢物感应。在这里,我们利用单分子 FRET 分析了第一个新生结构(5-́茎环)在转录延伸过程中展开形成配体结合合格结构的转变过程。我们的研究结果表明,结构转换以相对突然的方式发生,即在 1-2 个核苷酸窗口内。此外,我们还观察到了高度动态的结构交换,这表明核糖开关转录本对新生共存结构进行了快速取样。我们还观察到,在延伸过程中,RNAP的存在稳定了5́茎环,这与RNAP与5́茎环相互作用是一致的。我们的研究强调了早期折叠茎环结构在参与遗传调控的复杂 RNA 分子的共转录形成过程中的作用。
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来源期刊
Journal of Molecular Biology
Journal of Molecular Biology 生物-生化与分子生物学
CiteScore
11.30
自引率
1.80%
发文量
412
审稿时长
28 days
期刊介绍: Journal of Molecular Biology (JMB) provides high quality, comprehensive and broad coverage in all areas of molecular biology. The journal publishes original scientific research papers that provide mechanistic and functional insights and report a significant advance to the field. The journal encourages the submission of multidisciplinary studies that use complementary experimental and computational approaches to address challenging biological questions. Research areas include but are not limited to: Biomolecular interactions, signaling networks, systems biology; Cell cycle, cell growth, cell differentiation; Cell death, autophagy; Cell signaling and regulation; Chemical biology; Computational biology, in combination with experimental studies; DNA replication, repair, and recombination; Development, regenerative biology, mechanistic and functional studies of stem cells; Epigenetics, chromatin structure and function; Gene expression; Membrane processes, cell surface proteins and cell-cell interactions; Methodological advances, both experimental and theoretical, including databases; Microbiology, virology, and interactions with the host or environment; Microbiota mechanistic and functional studies; Nuclear organization; Post-translational modifications, proteomics; Processing and function of biologically important macromolecules and complexes; Molecular basis of disease; RNA processing, structure and functions of non-coding RNAs, transcription; Sorting, spatiotemporal organization, trafficking; Structural biology; Synthetic biology; Translation, protein folding, chaperones, protein degradation and quality control.
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